Self-assembled monolayers formed by the chemisorption of alkanethiols on gold are likely to now be the most intensively characterized synthetic organic monolayers prepared to date. See, Ulman, AN INTRODUCTION TO ULTRATHIN ORGANIC FILMS: FROM LANGMUIR-BLODGETT TO SELF ASSEMBLY, Academic Press, San Diego, 1991; Dubois el al., Annu. Rev. Phys. Chem., 43: 437 (1992). These monolayers form spontaneously during immersion of evaporated films of gold in solutions of alkanethiols as a result of chemisorption of sulfur on the (111) textured surface of the films. The molecules self-organize into a commensurate 3.times.3R30.degree. lattice on the surface of the Au(111). See, Porter, J. Am. Chem. Soc., 109: 3559 (1987); Camillone III, et al., Chem. Phys., 98: 3503 (1993); Fenter et al., Science, 266: 1216 (1994); 20; Chidsey et al., Langmuir, 6: 682 (1990); Sun etal., Thin Solid Films, 242: 106 (1994). For monolayers formed from CH.sub.3 (CH.sub.2).sub.n SH, n&gt;9, at least, the aliphatic chains of the monolayers are extended in the all-trans conformation and tilted approximately 30.degree. from the normal of the surface. Because the spacing between sulfur groups on the 3.times.3R30.degree. lattice is, on average, 4.9 .ANG., whereas the van der Waals diameter of an aliphatic chains is only .about.4 .ANG., the aliphatic chains within these SAMs tilt from the normal so as to come into van der Waals contact and thereby maximize their cohesive dispersive interactions. Studies of the lateral structure within monolayers using X-ray diffraction reveal the existence of domains of size .about.100 .ANG., where each domain has one of six different tilt directions relative to the Au(111) face. See, Fenter et al., Science, 266: 1216 (1994). Recent studies have shown the existence a c(4.times.2) superlattice, the cause of which remains unresolved.
Self-assembled monolayers formed with .omega.-substituted alkanethiols on the surface of gold have been used as model surfaces in a number of past studies of the interactions of proteins with surfaces (Spinke et al., Langumuir, 9: 1821 (1993); Willner et al., J. Am. Chem. Soc., 114: 10965 (1992); Song et al., J. Phys, Chem., 97: 6564 (1993); Mrksich et al., J. Am. Chem. Soc., 117: 12009 (1995)). For example, multilayer systems based on biotinylated alkanethiols and streptavidin have been used in schemes for the immobilization of Fab fragments of antibodies on surfaces (Spinke et al, Langumuir, 9: 1821 (1993)), and SAMs formed from NHS-activated disulfides have been used to form enzyme-based electrodes by covalent immobilization of glutathione reductase (Willner et al., J. Am. Chem. Soc., 114: 10965 (1992)). Cytochrome c, when adsorbed to SAMs formed from mercaptoundecanoic acid, has also been shown to be active and to possess a formal potential nearly identical to that of cytochrome c bound to physiological membranes (Song et al., J. Phys, Chem., 97: 6564 (1993)).
Whereas, investigations such as those described above have firmly established the use of SAMs for studies of specific interactions between proteins and surfaces, mixed SAMs formed from hydrophobic (methyl-terminated) and hydrophilic (hydroxyl-, oligo(ethylene glycol)-terminated) alkanethiols have also been used as model surfaces in studies of non-specific adsorption of proteins onto surfaces. Whitesides and coworkers, for example, have reported a study of the non-specific adsorption of fibrinogen, lysozyme, pyruvate kinase and RNAse to mixed SAMs (Prime et al., J. Am. Chem. Soc., 115: 10714 (1993); Prime et al., Science, 252: 1164 (1991)). By using ellipsometry, SAMs formed from oligo(ethylene glycol)-terminated alkanethiols were shown to resist irreversible adsorption of these proteins.
Surfaces prepared by the chemisorption of organosulflur compounds on evaporated films of gold are not limited to the alkanethiols. Self-assembled monolayers formed from perfluorinated organosulfur compounds have also been reported. See, Lenk et al., Langmuir, 10: 4610(1994); Drawhorn et al., J. Phys. Chem., 99: 16511 (1995). These surfaces, too, can be highly ordered, although, interestingly, the origin of the order within the monolayer is largely intramolecular and contrasts, therefore, to monolayers formed from alkanethiols (where the order largely reflects the cohesive intermolecular dispersion force). Steric interactions between adjacent fluorine atoms of a perfluorinated chain cause the chain to twists itself into a rigid, helical conformation. That is, an isolated perfluoro chain is stiff, as compared to an aliphatic chain. Because perfluorinated chains have larger cross-sectional areas than alkanethiols, monolayers formed on gold from perfluorinated thiols are not tilted from the normal to the same degree as alkanethiols. See, Drawhorn et al., J. Phys. Chem., 99: 16511 (1995). Estimates by IR studies place the tilt of the perfluorinated chains at 0.about.10.degree.. Because perfluorinated chains within SAMs on Au(111) are not tilted to the same degree as the alkanethiols, their surfaces are not expected to possess domains formed from regions of monolayer with different tilt directions (as occurs with monolayers formed from alkanethiols).
Past studies of the interactions of proteins and SAMs formed from alkanethiols on gold have used either planar surfaces prepared by electron-beam or thermal deposition of gold. See, for example, Prime et al., Science, 252: 1164 (1991); Prime et al., Science, 252: 1164 (1991) or highly curved surfaces formed by using colloidal gold (Brust et al., J. Chem. Soc. Chem. Commun., 1994: 801 (1994); Johnson et al., Langmuir, 13: 51 (1997); Green et al., J. Phys. Chem., 101: 2663 (1997); Natan et al., Anal. Chem., 69: 471 (1997); Grabar et al., Langmuir, 12: 2353 (1996); Hancock, HIGH PERFORMANCE LIQUID CHROMATOGRAPHY IN BIOTECHNOLOGY, Wiley, New York (1990)).
Whereas planar interfaces prepared by the evaporation of gold have surface areas that are too small to be generally useful for biological assays based on measurements of bulk concentrations of analytes, colloidal particles (.about.1-10 nm in size) are not large enough to pack (unsupported) in columns through which reagents can be readily passed. Due to the expense of the materials involved, the use of larger gold particles is not practical.
Various electroless plating techniques have been used to coat particles with layers of various metals. For example, Barder et al., U.S. Pat. No. 5,196,267, discloses silica microspheres having a diameter of about 0.1 .mu.m to about 10 .mu.m provided with a thin surface layer of a metal. The silica particles are contacted with an aqueous or alcoholic solution of a metal compound to deposit the surface layer. Moreover, Alexander et al., U.S. Pat. No. 4,944,985 discloses a process for the electroless plating of easily reducible metals onto ultrafine, usually inert particles. Neither of these references teaches forming an organic layer on the metal film.
Self assembled monolayers have been constructed on silica particles by a number of known methods, however, silica gel particles are a less than ideal substrate for a number of reasons.
First, the surface of silica is formed from silanol (Si--OH) and siloxane (Si--O--Si groups). The surface concentration of silanol groups typically ranges between 1-3 .mu.mol/m.sup.2 (Welsch et al., J. Chromatogr., 506: 97 (1990), Kohler et al., J. Chromatogr., 352: 275 (1986)). Fully hydroxylated silica can have an areal density of silanol groups as high as 8 .mu.mol/m.sup.2 (Tuel et al., Langmuir, 6: 770 (1990)). Because some fraction of these silanol groups are deprotonated at physiological pHs, the surface of silica gel presents a net negative charge under solution conditions that are not highly acidic. Electrostatic interactions between these surface-bound charges and proteins are one principal cause of non-specific adsorption of proteins onto silica gel (Hancock, HIGH PERFORMANCE LIQUID CHROMATOGRAPHY IN BIOTECHNOLOGY, Wiley, New York, 1990).
The second disadvantage of silica gel-based supports is that chemical modification of the surface of silica gel generally requires the use of silane-based chemistries (Sagiv, J. Am. Chem. Soc., 102: 92 (1980)). Although monomolecular layers of alkyltrichlorosilanes have been reported to form on silicon oxide surfaces (Xia et al., J. Am. Chem. Soc., 117: 9576 (1995)), reproducible preparation of such monolayers is a substantial challenge. For example, Netzer and coworkers have reported surface coverages of vinyl-terminated alkyltrichlorosilane to be approximately 63% of that of a densely-packed monolayer (Netzer et al., This Solid Films, 100: 67 (1983)). Also, Sagiv and coworkers have suggested that incomplete monolayers formed from alkyltrichlorosilanes have a heterogeneous island-like structure (Maoz et al., J. Colloid and Interf Sci., 100: 465 (1984); Cohen et al., J. Chem Phys., 90: 3054 (1986)). In contrast, Whitesides and coworkers have concluded that incomplete monolayers formed from alkyltrichlorosilanes are homogenous and disordered. (Wasserman et al., J. Am. Chem. Soc., 111: 5852 (1989); Tidswell et al, Phys. Rev. B., 41: 1111 (1990) and Ulman, AN INTRODUCTION TO ULTRATHIN ORGANIC FILMS: FROM LANGMUIR-BLODGETT TO SELF-ASSEMBLY, Academic Press, Boston, 1991).
Thus, a method of preparing a particulate material having a substantially homogeneous, easily assembled organic layer that does not adventitiously and/or nonspecifically bind charged species would represent a significant advance in the art. Surprisingly, the present invention provides such particles, and methods of preparing and using these particles.